Thyroid hormone (TH) is a key regulator of metabolic homeostasis in humans. It also plays a fundamental role in human development. Thus, an elegant feedback system has developed to tightly control circulating TH levels within a tight range. Central to this control is the negative regulation of thyrotropin-releasing hormone (TRH) gene expression in the paraventricular nucleus of the hypothalamus (PVH) by TH. In addition to TH, TRH is also regulated by leptin and melanocortin signaling pathways such that fasting represses TRH expression. However, the molecular mechanism by which TRH is negatively regulated by both TH and fasting is not known. On positively regulated genes TH interacts with thyroid hormone receptor isoforms (TRs) on TH response elements (TREs) to initially relieve repression caused by the unliganded receptor's ability to recruit a corepressor complex. In addition to relieving repression, TH further activates gene expression by allowing the TR to recruit a cast of coactivators which act as histone-modifying enzymes and enhance transcription. While corepressors and coactivators are presumed to play a role in negative regulation no in vivo model system has been developed to test there role. Furthermore, the identification of true negative TREs has remained elusive. Thus, understanding the regulation of TRH gene expression provides an ideal model to discern the mechanisms governing negative regulation by TH. In addition, new insight will be garnered in the cross-talk that must exist between leptin and TH signaling in the TH neuron. In this proposal we will use a number of new genetic models to allow us to determine the molecular mechanism governing the regulation of TRH gene expression. Aim 1 will employ novel mouse models to discern the role of coregulators in negative regulation of TRH by TH. Aim 2 will utilize a novel transgenic mouse which contains a TRH BAG that the targets the PVN and is regulated by TH. Manipulation of this BAG will allow for the identification for the first time of a negative TRE. Aim 3 will focus on both cross-talk between the leptin and TH pathways within the TRH neuron and also understanding the exact mechanism by which leptin and/or downstream pathways regulate TRH expression. Completion of these Aims will shed new light on how metabolic pathwys target gene expression to preserve metabolic homeostasis. This should allow for a better understanding of physiologic adaptation to disease.